Impact of Microscopic and Submicroscopic Parasitemia During Pregnancy on Placental Malaria in a High-Transmission Setting in Uganda.

BACKGROUND: Placental malaria is a major cause of adverse birth outcomes. However, data are limited on the relationships between longitudinal measures of parasitemia during pregnancy and placental malaria. METHODS: Data came from 637 women enrolled in a randomized controlled trial of intermittent preventive treatment of malaria in pregnancy (IPTp) from Uganda. Plasmodium falciparum parasitemia was assessed using microscopy and ultrasensitive quantitative PCR at intervals of 28 days from 12 to 20 weeks gestation through delivery. Multivariate analysis was used to measure associations between characteristics of parasitemia during pregnancy and the risk of placental malaria based on histopathology. RESULTS: Overall risk of placental malaria was 44.6%. None of the 34 women without parasitemia detected during pregnancy had evidence of placental malaria. Increasing proportion of interval assessments with parasitemia and higher parasite densities were independently associated with an increased risk of placental malaria. Higher gravidity and more effective IPTp were associated with a decreased risk of placental malaria. Women with parasitemia only detected before the third trimester still had an increased risk of placental malaria. CONCLUSIONS: The frequency, density, and timing of parasitemia are all important risk factors for placental malaria. Interventions should target the prevention of all levels of parasitemia throughout pregnancy

The prevalence of histologic acute chorioamnionitis among HIV infected pregnant women in Uganda and its association with adverse birth outcomes.

BACKGROUND: Preterm birth (PTB) is a leading cause of neonatal mortality and longer-term morbidity. Acute chorioamnionitis (ACA) is a common cause of PTB, however, there are limited data on the prevalence of ACA and its association with PTB in resource limited settings. METHODS: Data and samples came from a clinical trial evaluating novel strategies for the prevention of malaria in HIV infected pregnant women in Uganda. Women were enrolled between 12-28 weeks of gestation and followed through delivery. For each placenta delivered, three placental tissue types (membrane roll, umbilical cord and chorionic plate/villous parenchyma) were collected. Slides were assessed for diagnosis of maternal and fetal ACA by microscopic evaluation of neutrophilic infiltration using a standardized grading scale. The primary outcomes were PTB (<37 weeks), low birth weight (LBW, <2500 grams), and small-for-gestational age (SGA, birth weight <10th percentile for age). Univariate and multivariate logistic regression were used to estimate associations between 1) maternal characteristics (age, education, wealth, gravidity, gestational age at enrollment, placental malaria, anti-malarial prophylaxis treatment regimen, HIV disease parameters) and ACA, and 2) associations between ACA and adverse birth outcomes. FINDINGS: A total of 193 placentas were included in the analysis. The prevalence of maternal and fetal ACA was 44.5% and 28.0%, respectively. HIV infected women between 28-43 years of age had a higher risk of maternal ACA compared to those between 17-21 years of age (50.9% vs. 19.1%; aOR = 4.00 (1.10-14.5), p = 0.04) and the diagnosis of severe maternal ACA was associated with a significantly higher risk of PTB (28.6% vs. 6.0%; aOR = 6.04 (1.87-19.5), p = 0.003), LBW (33.3% vs. 9.4%; aOR = 4.86 (1.65-14.3); p = 0.004), and SGA (28.6% vs. 10.1%; aOR = 3.70 (1.20-11.4), p = 0.02). No maternal characteristics were significantly associated with fetal ACA and the diagnosis of fetal ACA was not associated with adverse birth outcomes. CONCLUSIONS: Histological evidence of severe maternal ACA was associated with an increased risk of PTB, LBW, and SGA in HIV infected, pregnant Ugandan women

HRP2 and pLDH-Based Rapid Diagnostic Tests, Expert Microscopy, and PCR for Detection of Malaria Infection during Pregnancy and at Delivery in Areas of Varied Transmission: A Prospective Cohort Study in Burkina Faso and Uganda.

BACKGROUND: Intermittent screening and treatment (IST) of malaria during pregnancy has been proposed as an alternative to intermittent preventive treatment in pregnancy (IPTp), where IPTp is failing due to drug resistance. However, the antenatal parasitaemias are frequently very low, and the most appropriate screening test for IST has not been defined. METHODOLOGY/PRINCIPAL FINDINGS: We conducted a multi-center prospective study of 990 HIV-uninfected women attending ANC in two different malaria transmission settings at Tororo District Hospital, eastern Uganda and Colsama Health Center in western Burkina Faso. Women were enrolled in the study in the second or third trimester of pregnancy and followed to delivery, generating 2,597 blood samples for analysis. Screening tests included rapid diagnostic tests (RDTs) targeting histidine-rich protein 2 (HRP2) and parasite lactate dehydrogenase (pLDH) and microscopy, compared to nPCR as a reference standard. At enrolment, the proportion of pregnant women who were positive for P. falciparum by HRP2/pan pLDH RDT, Pf pLDH/pan pLDH RDT, microscopy and PCR was 38%, 29%, 36% and 44% in Uganda and 21%, 16%, 15% and 35% in Burkina Faso, respectively. All test positivity rates declined during follow-up. In comparison to PCR, the sensitivity of the HRP2/pan pLDH RDT, Pf pLDH/pan pLDH RDT and microscopy was 75.7%, 60.1% and 69.7% in Uganda, 55.8%, 42.6% and 55.8% in Burkina Faso respectively for all antenatal visits. Specificity was greater than 96% for all three tests. Comparison of accuracy using generalized estimating equation revealed that the HRP2- detecting RDT was the most accurate test in both settings. CONCLUSIONS/SIGNIFICANCE: The study suggests that HRP2-based RDTs are the most appropriate point-of-care test currently available for use during pregnancy especially for symptomatic women, but will still miss some PCR-positive women. The clinical significance of these very low density infections needs to be better defined

Relationships Between Measures of Malaria at Delivery and Adverse Birth Outcomes in a High-Transmission Area of Uganda.

BACKGROUND: Clinical trials of interventions for preventing malaria in pregnancy often use measures of malaria at delivery as their primary outcome. Although the objective of these interventions is to improve birth outcomes, data on associations between different measures of malaria at delivery and adverse birth outcomes are limited. METHODS: Data came from 637 Ugandan women enrolled in a randomized controlled trial of intermittent preventive treatment of malaria in pregnancy. Malaria at delivery was detected using peripheral and placental blood microscopy, placental blood loop-mediated isothermal amplification (LAMP), and placental histopathology. Multivariate analyses were used to estimate associations between measures of malaria at delivery and risks of low birth weight (LBW), small for gestational age (SGA), and preterm birth (PTB). RESULTS: Detection of malaria parasites by microscopy or LAMP was not associated with adverse birth outcomes. Presence of malaria pigment detected by histopathology in ?30% of high-powered fields was strongly associated with LBW (adjusted risk ratio [aRR] = 3.42, P = .02) and SGA (aRR = 4.24, P < .001) but not PTB (aRR = 0.88, P = .87). CONCLUSIONS: A semiquantitative classification system based on histopathologically detected malaria pigment provided the best surrogate measure of adverse birth outcomes in a high-transmission setting and should be considered for use in malaria in pregnancy intervention studies

The impact of gravidity, symptomatology and timing of infection on placental malaria.

BACKGROUND: Placental malaria is associated with increased risk of adverse perinatal outcomes. While primigravidity has been reported as a risk factor for placental malaria, little is known regarding the relationship between gravidity, symptomatology and timing of Plasmodium falciparum infection and the development of placental malaria. METHODS: The aim of this study was to investigate the relationship between the development of placental malaria and gravidity, timing of infection, and presence of symptoms. This is a secondary analysis of data from a double-blind randomized control trial of intermittent preventive therapy during pregnancy in Uganda. Women were enrolled from 12 to 20 weeks gestation and followed through delivery. Exposure to malaria parasites was defined as symptomatic (fever with positive blood smear) or asymptomatic (based on molecular detection of parasitaemia done routinely every 4 weeks). The primary outcome was placental malaria diagnosed by histopathology, placental blood smear, and/or placental blood loop-mediated isothermal amplification. Multivariate analyses were performed using logistic regression models. Subgroup analysis was performed based on the presence of symptomatic malaria, gravidity, and timing of infection. RESULTS: Of the 228 patients with documented maternal infection with malaria parasites during pregnancy, 101 (44.3%) had placental malaria. Primigravidity was strongly associated with placental malaria (aOR 8.90, 95% CI 4.34-18.2, p < 0.001), and each episode of malaria was associated with over a twofold increase in placental malaria (aOR 2.35, 95% CI 1.69-3.26, p < 0.001). Among multigravid women, the odds of placental malaria increased by 14% with each advancing week of gestation at first documented infection (aOR 1.14, 95% CI 1.02-1.27, p = 0.02). When stratified by the presence of symptoms, primigravidity was only associated with placental malaria in asymptomatic women, who had a 12-fold increase in the odds of placental malaria (aOR 12.19, 95% CI 5.23-28.43, p < 0.001). CONCLUSIONS: Total number of P. falciparum infections in pregnancy is a significant predictor of placental malaria. The importance of timing of infection on the development of placental malaria varies based on gravidity. In primigravidas, earlier asymptomatic infections were more frequently identified in those with placental malaria, whereas in multigravidas, parasitaemias detected later in gestation were associated with placental malaria. Earlier initiation of an effective intermittent preventive therapy may help to prevent placental malaria and improve birth outcomes, particularly in primigravid women

Infant sex modifies associations between placental malaria and risk of malaria in infancy.

BACKGROUND: Placental malaria (PM) has been associated with a higher risk of malaria during infancy. However, it is unclear whether this association is causal, and is modified by infant sex, and whether intermittent preventive treatment in pregnancy (IPTp) can reduce infant malaria by preventing PM. METHODS: Data from a birth cohort of 656 infants born to HIV-uninfected mothers randomised to IPTp with dihydroartemisinin-piperaquine (DP) or Sulfadoxine-pyrimethamine (SP) was analysed. PM was categorized as no PM, active PM (presence of parasites), mild-moderate past PM (> 0-20% high powered fields [HPFs] with pigment), or severe past PM (> 20% HPFs with pigment). The association between PM and incidence of malaria in infants stratified by infant sex was examined. Causal mediation analysis was used to test whether IPTp can impact infant malaria incidence via preventing PM. RESULTS: There were 1088 malaria episodes diagnosed among infants during 596.6 person years of follow-up. Compared to infants born to mothers with no PM, the incidence of malaria was higher among infants born to mothers with active PM (adjusted incidence rate ratio [aIRR] 1.30, 95% CI 1.00-1.71, p = 0.05) and those born to mothers with severe past PM (aIRR 1.28, 95% CI 0.89-1.83, p = 0.18), but the differences were not statistically significant. However, when stratifying by infant sex, compared to no PM, severe past PM was associated a higher malaria incidence in male (aIRR 2.17, 95% CI 1.45-3.25, p < 0.001), but not female infants (aIRR 0.74, 95% CI 0.46-1.20, p = 0.22). There were no significant associations between active PM or mild-moderate past PM and malaria incidence in male or female infants. Male infants born to mothers given IPTp with DP had significantly less malaria in infancy than males born to mothers given SP, and 89.7% of this effect was mediated through prevention of PM. CONCLUSION: PM may have more severe consequences for male infants, and interventions which reduce PM could mitigate these sex-specific adverse outcomes. More research is needed to better understand this sex-bias between PM and infant malaria risk. Trial registration ClinicalTrials.gov, NCT02793622. Registered 8 June 2016, https://clinicaltrials.gov/ct2/show/NCT02793622

Reductions in malaria in pregnancy and adverse birth outcomes following indoor residual spraying of insecticide in Uganda.

BACKGROUND: Indoor residual spraying of insecticide (IRS) is a key intervention for reducing the burden of malaria in Africa. However, data on the impact of IRS on malaria in pregnancy and birth outcomes is limited. METHODS: An observational study was conducted within a trial of intermittent preventive therapy during pregnancy in Tororo, Uganda. Women were enrolled at 12-20 weeks of gestation between June and October 2014, provided with insecticide-treated bed nets, and followed through delivery. From December 2014 to February 2015, carbamate-containing IRS was implemented in Tororo district for the first time. Exact spray dates were collected for each household. The exposure of interest was the proportion of time during a woman's pregnancy under protection of IRS, with three categories of protection defined: no IRS protection, >0-20 % IRS protection, and 20-43 % IRS protection. Outcomes assessed included malaria incidence and parasite prevalence during pregnancy, placental malaria, low birth weight (LBW), pre-term delivery, and fetal/neonatal deaths. RESULTS: Of 289 women followed, 134 had no IRS protection during pregnancy, 90 had >0-20 % IRS protection, and 65 had >20-43 % protection. During pregnancy, malaria incidence (0.49 vs 0.10 episodes ppy, P = 0.02) and parasite prevalence (20.0 vs 8.9 %, P 20-43 % IRS protection (1.6 %, P = 0.006). Compared to women with no IRS protection, those with >20-43 % IRS protection had a lower risk of LBW (20.9 vs 3.1 %, P = 0.002), pre-term birth (17.2 vs 1.5 %, P = 0.006), and fetal/neonatal deaths (7.5 vs 0 %, P = 0.03). CONCLUSION: In this setting, IRS was temporally associated with lower malaria parasite prevalence during pregnancy and at delivery, and improved birth outcomes. IRS may represent an important tool for combating malaria in pregnancy and for improving birth outcomes in malaria-endemic settings. Trial Registration Current Controlled Trials Identifier NCT02163447

Relationships between infection with Plasmodium falciparum during pregnancy, measures of placental malaria, and adverse birth outcomes.

BACKGROUND: Malaria in pregnancy has been associated with maternal morbidity, placental malaria, and adverse birth outcomes. However, data are limited on the relationships between longitudinal measures of malaria during pregnancy, measures of placental malaria, and birth outcomes. METHODS: This is a nested observational study of data from a randomized controlled trial of intermittent preventive therapy during pregnancy among 282 participants with assessment of placental malaria and delivery outcomes. HIV-uninfected pregnant women were enrolled at 12-20 weeks of gestation. Symptomatic malaria during pregnancy was measured using passive surveillance and monthly detection of asymptomatic parasitaemia using loop-mediated isothermal amplification (LAMP). Placental malaria was defined as either the presence of parasites in placental blood by microscopy, detection of parasites in placental blood by LAMP, or histopathologic evidence of parasites or pigment. Adverse birth outcomes assessed included low birth weight (LBW), preterm birth (PTB), and small for gestational age (SGA) infants. RESULTS: The 282 women were divided into three groups representing increasing malaria burden during pregnancy. Fifty-two (18.4%) had no episodes of symptomatic malaria or asymptomatic parasitaemia during the pregnancy, 157 (55.7%) had low malaria burden (0-1 episodes of symptomatic malaria and < 50% of samples LAMP+), and 73 (25.9%) had high malaria burden during pregnancy (≥ 2 episodes of symptomatic malaria or ≥ 50% of samples LAMP+). Women with high malaria burden had increased risks of placental malaria by blood microscopy and LAMP [aRR 14.2 (1.80-111.6) and 4.06 (1.73-9.51), respectively], compared to the other two groups combined. Compared with women with no malaria exposure during pregnancy, the risk of placental malaria by histopathology was higher among low and high burden groups [aRR = 3.27 (1.32-8.12) and aRR = 7.07 (2.84-17.6), respectively]. Detection of placental parasites by any method was significantly associated with PTB [aRR 5.64 (1.46-21.8)], and with a trend towards increased risk for LBW and SGA irrespective of the level of malaria burden during pregnancy. CONCLUSION: Higher malaria burden during pregnancy was associated with placental malaria and together with the detection of parasites in the placenta were associated with increased risk for adverse birth outcomes. Trial Registration Current Controlled Trials Identifier NCT02163447

Dihydroartemisinin-Piperaquine for the Prevention of Malaria in Pregnancy.

BACKGROUND: Intermittent treatment with sulfadoxine-pyrimethamine is widely recommended for the prevention of malaria in pregnant women in Africa. However, with the spread of resistance to sulfadoxine-pyrimethamine, new interventions are needed. METHODS: We conducted a double-blind, randomized, controlled trial involving 300 human immunodeficiency virus (HIV)-uninfected pregnant adolescents or women in Uganda, where sulfadoxine-pyrimethamine resistance is widespread. We randomly assigned participants to a sulfadoxine-pyrimethamine regimen (106 participants), a three-dose dihydroartemisinin-piperaquine regimen (94 participants), or a monthly dihydroartemisinin-piperaquine regimen (100 participants). The primary outcome was the prevalence of histopathologically confirmed placental malaria. RESULTS: The prevalence of histopathologically confirmed placental malaria was significantly higher in the sulfadoxine-pyrimethamine group (50.0%) than in the three-dose dihydroartemisinin-piperaquine group (34.1%, P=0.03) or the monthly dihydroartemisinin-piperaquine group (27.1%, P=0.001). The prevalence of a composite adverse birth outcome was lower in the monthly dihydroartemisinin-piperaquine group (9.2%) than in the sulfadoxine-pyrimethamine group (18.6%, P=0.05) or the three-dose dihydroartemisinin-piperaquine group (21.3%, P=0.02). During pregnancy, the incidence of symptomatic malaria was significantly higher in the sulfadoxine-pyrimethamine group (41 episodes over 43.0 person-years at risk) than in the three-dose dihydroartemisinin-piperaquine group (12 episodes over 38.2 person-years at risk, P=0.001) or the monthly dihydroartemisinin-piperaquine group (0 episodes over 42.3 person-years at risk, P<0.001), as was the prevalence of parasitemia (40.5% in the sulfadoxine-pyrimethamine group vs. 16.6% in the three-dose dihydroartemisinin-piperaquine group [P<0.001] and 5.2% in the monthly dihydroartemisinin-piperaquine group [P<0.001]). In each treatment group, the risk of vomiting after administration of any dose of the study agents was less than 0.4%, and there were no significant differences among the groups in the risk of adverse events. CONCLUSIONS: The burden of malaria in pregnancy was significantly lower among adolescent girls or women who received intermittent preventive treatment with dihydroartemisinin-piperaquine than among those who received sulfadoxine-pyrimethamine, and monthly treatment with dihydroartemisinin-piperaquine was superior to three-dose dihydroartemisinin-piperaquine with regard to several outcomes. (Funded by the Eunice Kennedy Shriver National Institute of Child Health and Human Development; ClinicalTrials.gov number, NCT02163447.)

HRP2 and pLDH-based rapid diagnostic tests, expert microscopy, and PCR for detection of malaria infection during pregnancy and at delivery in areas of varied transmission: A prospective cohort study in Burkina Faso and Uganda

Background Intermittent screening and treatment (IST) of malaria during pregnancy has been proposed as an alternative to intermittent preventive treatment in pregnancy (IPTp), where IPTp is failing due to drug resistance. However, the antenatal parasitaemias are frequently very low, and the most appropriate screening test for IST has not been defined. Methodology/Principal Findings We conducted a multi-center prospective study of 990 HIV-uninfected women attending ANC in two different malaria transmission settings at Tororo District Hospital, eastern Uganda and Colsama Health Center in western Burkina Faso. Women were enrolled in the study in the second or third trimester of pregnancy and followed to delivery, generating 2,597 blood samples for analysis. Screening tests included rapid diagnostic tests (RDTs) targeting histidine-rich protein 2 (HRP2) and parasite lactate dehydrogenase (pLDH) and microscopy, compared to nPCR as a reference standard. At enrolment , the proportion of pregnant women who were positive for P. falciparum by HRP2/pan pLDH RDT, Pf pLDH/pan pLDH RDT, microscopy and PCR was 38%, 29%, 36% and 44% in Uganda and 21%, 16%, 15% and 35% in Bukina Faso, respectively. All test positivity ratesdeclined during follow-up. In comparison to PCR, the sensitivity of the HRP2/pan pLDH RDT, Pf pLDH/pan pLDH RDT and microscopy was 75.7 %, 60.1% and 69.7% in Uganda, 55.8%, 42.6% and 55.8% in Burkina Faso respectively for all antenatal visits. Specificity was greater than 96% for all three tests. Comparison of accuracy using generalized estimating equation revealed that the HRP2- detecting RDT was the most accurate test in both settings. Conclusions/Significance The study suggests that HRP2-based RDTs are the most appropriate point-of-care test currently available for use during pregnancy especially for symptomatic women, but will still miss some PCR-positive women. The clinical significance of these very low density infections needs to be better defined